scholarly journals Assessment of SARS-CoV-2 infectivity of upper respiratory specimens from COVID-19 patients by virus isolation using VeroE6/TMPRSS2 cells

2021 ◽  
Vol 8 (1) ◽  
pp. e000830
Author(s):  
Souichi Yamada ◽  
Shuetsu Fukushi ◽  
Hitomi Kinoshita ◽  
Makoto Ohnishi ◽  
Tadaki Suzuki ◽  
...  
Keyword(s):  
Virus Isolation ◽  
Laboratory Diagnostics ◽  
Viral Isolation ◽  
Viral Loads ◽  
A Cell ◽  
Blind Passage ◽  
Upper Respiratory ◽  
Novel Coronavirus ◽  
Respiratory Specimens

BackgroundAn outbreak of novel coronavirus (SARS-CoV-2)-associated respiratory infectious diseases (COVID-19) emerged in 2019 and has spread rapidly in humans around the world. The demonstration of in vitro infectiousness of respiratory specimens is an informative surrogate for SARS-CoV-2 transmission from patients with COVID-19; accordingly, viral isolation assays in cell culture are an important aspect of laboratory diagnostics for COVID-19.MethodsWe developed a simple and rapid protocol for isolating SARS-CoV-2 from respiratory specimens using VeroE6/TMPRSS2 cells, a cell line that is highly susceptible to the virus. We also investigated a correlation between isolation of SARS-CoV-2 and viral load detected by real-time RT-PCR (rRT-PCR) using N2 primer/probe set that has been developed for testing of COVID-19 in Japan.ResultsThe SARS-CoV-2 isolation protocol did not require blind passage of inoculated cells and yielded the results of viral isolation within 7 days after inoculation. Specimens with cycle threshold (Ct) values of <20.2, determined by rRT-PCR, were predicted to be isolation-positive. On the other hand, 6.9% of specimens with Ct values >35 were virus isolation-positive, indicating that low viral loads (high Ct values) in upper respiratory specimens do not always indicate no risk of containing transmissible virus.ConclusionIn combination with rRT-PCR, the SARS-CoV-2 isolation protocol provides a means for assessing the potential risk of transmissible virus in upper respiratory specimens.

Antiviral Properties of Food Plants Could Help to Reduce Contagion and Severity in SARS-CoV-2 Infections

2021 ◽  
Vol 07 ◽  
Author(s):  
Betina Cardoso
Keyword(s):  
Respiratory Tract ◽  
Literature Search ◽  
Food Plants ◽  
Upper Respiratory Tract ◽  
Site Specific ◽  
In Vivo Experiments ◽  
Upper Respiratory ◽  
Novel Coronavirus

Introduction: The importance of an immediate tool to help patients and prevent viral diffusion of new pneumonia caused by 2019 novel coronavirus (2019-nCoV or SARS-CoV-2) that causes the disease COVID-19 becomes evident. Recent articles have reported on body site-specific SARS-CoV-2 infection, showing very active replication in the throat and upper respiratory tract when symptoms were still mild, and thus being efficient in viral transmission in sputum. Material and Methods: An alternative that may be feasible is to resort to scientific studies that demonstrate the antiviral potential of medicinal plants species through in-vitro and in-vivo experiments to alleviate symptoms and prevent the spread of contagion. A literature search in Scopus and PubMed on herbs and foods with antiviral properties was performed. Results: Herbs and foods with demonstrated antiviral potential have been identified, which could limit SARS-CoV-2 spreading by interfering on ACE2 protein on infection sites. The analysis of transdisciplinary knowledge allows us to connect previous research on the action of common plants and foods on viruses to limit the replication of SARS-CoV-2 in the throat and upper respiratory tract. Conclusions: Herbs and foods with demonstrated antiviral potential have been identified, which could limit SARS-CoV-2 spreading by interfering on ACE2 protein on infection sites. The analysis of transdisciplinary knowledge allows us to connect previous research on the action of common plants and foods on viruses to limit the replication of SARS-CoV-2 in the throat and upper respiratory tract.

SARS-CoV-2 Variants of Concern B.1.1.7 and B.1.351 Are Not Associated With Higher Viral Loads in Upper Respiratory Specimens

2021 ◽  
Author(s):  
Chun Huai Luo ◽  
Adannaya Amadi ◽  
C. Paul Morris ◽  
Matthew Schwartz ◽  
Eili Y. Klein ◽  
...  
Keyword(s):  
Viral Loads ◽  
Upper Respiratory ◽  
Respiratory Specimens

scholarly journals Impact of the Nucleic Acid Extraction Method and the RT-qPCR Assay on SARS-CoV-2 Detection in Low-Viral Samples

Diagnostics ◽  
2021 ◽  
Vol 11 (12) ◽  
pp. 2247
Author(s):  
Magdalena Komiazyk ◽  
Jarosław Walory ◽  
Aleksandra Kozinska ◽  
Izabela Wasko ◽  
Anna Baraniak
Keyword(s):  
Rna Extraction ◽  
Extraction Methods ◽  
Etiologic Agent ◽  
Diagnostic Methods ◽  
Clinical Samples ◽  
Acid Extraction ◽  
Upper Respiratory Tract ◽  
Viral Loads ◽  
Upper Respiratory ◽  
Novel Coronavirus

COVID-19 was initially reported in China at the end of 2019 and soon thereafter, in March 2020, the WHO declared it a pandemic. Until October 2021, over 240 million COVID-19 cases were recorded, with 4.9 mln deaths. In order to stop the spread of this disease, it is crucial to monitor and detect any infected person. The etiologic agent of COVID-19 is a novel coronavirus called SARS-CoV-2. The gold standard for the detection of the virus is the RT-qPCR method. This study evaluated two RNA extraction methods and four commercial RT-qPCR assays routinely used in diagnostic laboratories for detecting SARS-CoV-2 in human specimens from the upper respiratory tract. We analyzed a panel of 70 clinical samples with varying RNA loads. Our study demonstrated the significant impact of the diagnostic methods selected by the laboratory on the SARS-CoV-2 detection in clinical specimens with low viral loads.

scholarly journals A sprayable Acid-Oxidizing solution containing hypochlorous acid (AOS2020) efficiently and safely inactivates SARS-Cov-2: a new potential solution for upper respiratory tract hygiene

2021 ◽  
Author(s):  
Nadia Giarratana ◽  
Balan Rajan ◽  
Kannan Kamala ◽  
Michelle Mendenhall ◽  
Giorgio Reiner
Keyword(s):  
Respiratory Tract ◽  
Oral Mucosa ◽  
Hypochlorous Acid ◽  
Tolerability Profile ◽  
Upper Respiratory Tract ◽  
Potential Solution ◽  
Viral Loads ◽  
Upper Respiratory ◽  
Virucidal Efficacy

Abstract Introduction To eliminate the COVID-19 pandemic, the transmission of the virus SARS-CoV-2 among the population needs to be blocked and/or at least reduced. Upper respiratory tract viral loads are highest in the early stages of the disease, and high loads are associated with higher mortality rates. This study aims to evaluate the virucidal efficacy of AOS2020, a novel sprayable Acid-Oxidizing solution containing pure and stable hypochlorous acid (HClO), on human coronavirus SARS-Cov-2 in vitro, and the tolerability profile on nasal and oral mucosa suggesting to be a potential solution for upper respiratory hygiene. Method Virucidal assays and intranasal and oral irritation tests were undertaken in accordance with relevant national and international guidance and methods. Results In pre-clinical tests, the AOS2020, showed > 99.8% virucidal efficacy in < 1 min against SARS-Cov-2. The safety profile testing on both the nasal and oral mucosa indicates that AOS2020 is non-irritant. Conclusion These initial results indicate that this product has the potential treatment to reduce viral load in the upper respiratory tract.

scholarly journals Comparing immunoassays for SARS-Coronavirus-2 antibody detection in patients with and without laboratory-confirmed SARS-Coronavirus-2 infection

2021 ◽  
Author(s):  
Karoline Leuzinger ◽  
Michael Osthoff ◽  
Sarah Dräger ◽  
Hans Pargger ◽  
Martin Siegemund ◽  
...  
Keyword(s):  
Antibody Detection ◽  
Upper Respiratory Tract ◽  
Sars Coronavirus ◽  
Cross Sectional ◽  
Viral Loads ◽  
Reactive Protein ◽  
Antibody Assays ◽  
Quantitative Results ◽  
Upper Respiratory ◽  
Novel Coronavirus

Background. Commercially available SARS-CoV-2-directed antibody assays may assist in diagnosing past exposure to SARS-CoV-2 antigens. Methods. We cross-compared eight immunoassays detecting antibodies against SARS-CoV-2 nucleocapsid(N)- or spike(S)-antigens in three cohorts consisting of 859 samples from 622 patients: (#1)EDI™-Novel-Coronavirus-COVID19, Epitope; (#2)RecomWell-SARS-CoV-2, Mikrogen; (#3)COVID19-ELISA, VirCell; (#4)Elecsys-Anti-SARS-CoV-2-N, Roche; (#5)LIAISON®-SARS-CoV-2-S1/S2, Diasorin; (#6)Anti-SARS-CoV-2-ELISA, EuroImmun; (#7)Elecsys-Anti-SARS-CoV-2-S, Roche; and (#8)LIAISON®-SARS-CoV-2-TrimericS, Diasorin. Results. In cross-sectional Cohort-1 (68 sera from 38 patients with documented SARS-CoV-2 infection), agreement between assays #1 to #6 ranged from 75% to 93%, whereby discordance mostly resulted from N-based assays #1 to #4. In cross-sectional Cohort-2 (510 sera from 510 patients; 56 documented, 454 unknown SARS-CoV-2 infection), assays #4 to #6 were analyzed further together with #7 and #8 revealing 94% concordance (44 [9%] positives and 485 [85%] negatives). Discordance was highest within 2 weeks after SARS-CoV-2/CoVID19 diagnosis and confirmed in the longitudinal Cohort-3 (281 sera from 74 CoVID19 patients), using assays #4, #6, #7 and #8. Sub-analysis of 20 (27%) initially seronegative Cohort-3 patients revealed assay-dependent 50% and 90% seroconversion rates after 8-11 days and 14-18 days, respectively. Increasing SARS-CoV-2 antibodies were significantly associated with declining levels of viral loads, lactate dehydrogenase, interleukin-6 and C-reactive protein and preceded clearance of SARS-CoV-2 detection in the upper respiratory tract by approximately 1 week. Conclusion. SARS-CoV-2 specific antibody assays show substantial agreement, but interpretation of qualitative and semi-quantitative results depends on the time elapsed post-diagnosis and the choice of viral antigen. Mounting of systemic SARS-CoV-2-specific antibodies may predict recovery from viral injury and clearance of mucosal replication.

Ultrastructure of melanocyte-keratinocyte interactions and pigment transfer in vitro

1978 ◽  
Vol 36 (2) ◽  
pp. 650-651
Author(s):  
Raul I. Garcia ◽  
Evelyn A. Flynn ◽  
George Szabo
Keyword(s):  
Direct Injection ◽  
Skin Pigmentation ◽  
Freeze Fracture ◽  
Pigment Granule ◽  
Time Lapse ◽  
Ultrastructural Level ◽  
Lanthanum Tracer ◽  
A Cell

Skin pigmentation in mammals involves the interaction of epidermal melanocytes and keratinocytes in the structural and functional unit known as the Epidermal Melanin Unit. Melanocytes(M) synthesize melanin within specialized membrane-bound organelles, the melanosome or pigment granule. These are subsequently transferred by way of M dendrites to keratinocytes(K) by a mechanism still to be clearly defined. Three different, though not necessarily mutually exclusive, mechanisms of melanosome transfer have been proposed: cytophagocytosis by K of M dendrite tips containing melanosomes, direct injection of melanosomes into the K cytoplasm through a cell-to-cell pore or communicating channel formed by localized fusion of M and K cell membranes, release of melanosomes into the extracellular space(ECS) by exocytosis followed by K uptake using conventional phagocytosis. Variability in methods of transfer has been noted both in vivo and in vitro and there is evidence in support of each transfer mechanism. We Have previously studied M-K interactions in vitro using time-lapse cinemicrography and in vivo at the ultrastructural level using lanthanum tracer and freeze-fracture.

Using FRET to Measure the Time it Takes for a Cell to Destroy a Virus

2019 ◽  
Author(s):  
Candace E. Benjamin ◽  
Zhuo Chen ◽  
Olivia Brohlin ◽  
Hamilton Lee ◽  
Stefanie Boyd ◽  
...  
Keyword(s):  
Cellular Uptake ◽  
Rhodamine B ◽  
Virus Like Particle ◽  
A Cell ◽  
Viral Nanoparticles ◽  
The Stability ◽  
Open Question ◽  
Viral Surface ◽  
Fret Pair

<div><div><div><p>The emergence of viral nanotechnology over the preceding two decades has created a number of intellectually captivating possible translational applications; however, the in vitro fate of the viral nanoparticles in cells remains an open question. Herein, we investigate the stability and lifetime of virus-like particle (VLP) Qβ - a representative and popular VLP for several applications - following cellular uptake. By exploiting the available functional handles on the viral surface, we have orthogonally installed the known FRET pair, FITC and Rhodamine B, to gain insight of the particle’s behavior in vitro. Based on these data, we believe VLPs undergo aggregation in addition to the anticipated proteolysis within a few hours of cellular uptake.</p></div></div></div>

Preincubation with Sn-complexes causes intensive intracellular retention of 99mTc in thyroid cells in vitro

2012 ◽  
Vol 51 (05) ◽  
pp. 179-185 ◽  
Author(s):  
M. Wendisch ◽  
D. Aurich ◽  
R. Runge ◽  
R. Freudenberg ◽  
J. Kotzerke ◽  
...  
Keyword(s):  
Cell Model ◽  
Low Energy ◽  
Thyroid Cells ◽  
Low Energy Electrons ◽  
A Cell ◽  
Vitro Model ◽  
Uptake Studies ◽  
Radiobiological Effects ◽  
Radioactivity Retention

SummaryTechnetium radiopharmaceuticals are well established in nuclear medicine. Besides its well-known gamma radiation, 99mTc emits an average of five Auger and internal conversion electrons per decay. The biological toxicity of these low-energy, high-LET (linear energy transfer) emissions is a controversial subject. One aim of this study was to estimate in a cell model how much 99mTc can be present in exposed cells and which radiobiological effects could be estimated in 99mTc-overloaded cells. Methods: Sodium iodine symporter (NIS)- positive thyroid cells were used. 99mTc-uptake studies were performed after preincubation with a non-radioactive (cold) stannous pyro - phosphate kit solution or as a standard 99mTc pyrophosphate kit preparation or with pure pertechnetate solution. Survival curves were analyzed from colony-forming assays. Results: Preincubation with stannous complexes causes irreversible intracellular radioactivity retention of 99mTc and is followed by further pertechnetate influx to an unexpectedly high 99mTc level. The uptake of 99mTc pertechnetate in NIS-positive cells can be modified using stannous pyrophosphate from 3–5% to >80%. The maximum possible cellular uptake of 99mTc was 90 Bq/cell. Compared with nearly pure extracellular irradiation from routine 99mTc complexes, cell survival was reduced by 3–4 orders of magnitude after preincubation with stannous pyrophosphate. Conclusions: Intra cellular 99mTc retention is related to reduced survival, which is most likely mediated by the emission of low-energy electrons. Our findings show that the described experiments constitute a simple and useful in vitro model for radiobiological investigations in a cell model.

Treatment Options in COVID-19: The Role of Bioavailable Antiviral Ribonucleoside Analog on NHC in vitro

2020 ◽  
Author(s):  
Lara Bittmann
Keyword(s):  
World Health Organization ◽  
Clinical Picture ◽  
Treatment Options ◽  
World Health ◽  
Wuhan City ◽  
The World ◽  
Novel Coronavirus ◽  
Health Organization

On December 31, 2019, WHO was informed of cases of pneumonia of unknown cause in Wuhan City, China. A novel coronavirus was identified as the cause by Chinese authorities on January 7, 2020 and was provisionally named "2019-nCoV". This new Coronavirus causes a clinical picture which has received now the name COVID-19. The virus has spread subsequently worldwide and was explained on the 11th of March, 2020 by the World Health Organization to the pandemic.

Analysis of the Dynamics of the Electric Capacitance of a Cell Monolayer at the Late Stages of Caco-2 cell Differentiation

2019 ◽  
Vol 35 (6) ◽  
pp. 87-90
Author(s):  
S.V. Nikulin ◽  
V.A. Petrov ◽  
D.A. Sakharov
Keyword(s):  
Impedance Spectroscopy ◽  
Cell Monolayer ◽  
Microfluidic Chips ◽  
Russian Science ◽  
Electric Capacitance ◽  
A Cell ◽  
Static Conditions ◽  
Late Stages

The real-time monitoring of electric capacitance (impedance spectroscopy) allowed obtaining evidence that structures which look like intestinal villi can be formed during the cultivation under static conditions as well as during the cultivation in microfluidic chips. It was shown in this work via transcriptome analysis that the Hh signaling pathway is involved in the formation of villus-like structures in vitro, which was previously shown for their formation in vivo. impedance spectroscopy, intestine, villi, electric capacitance, Hh The study was funded by the Russian Science Foundation (Project 16-19-10597).

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